CD34⁺CD38⁻ progenitor cells were seeded at a density of 1 cell per well in serum-depleted medium supplemented with Tpo in Terasaki plates. Shortly after plating (2 to 10 hours after seeding to allow cells to sediment), wells containing single cells were identified and all other wells were excluded from the experiment. After 116 hours of preincubation in the presence of Tpo, wells were reexamined to exclude wells containing more than 1 cell, ie, wells in which proliferation had occurred. A cocktail of cytokines (containing IL-1, IL-3, IL-6, G-CSF, GM-CSF, CSF-1, FL, KL, Epo, and Tpo) was added to wells containing a single cell directly after plating and where no proliferation had occurred during the first 116 hours of culture. Clonal growth was evaluated after an additional 14 days of incubation at 37°C and 5% CO₂ in air. Scoring criteria: 1, wells with 2 to 4 cells; 2, clones with 5 to 50 cells; 3, more than 50 cells but covering less than 10% of well; and 4, colonies covering more than 10% of well. The results represent the means (±SEM) of three individual experiments.